Lifelong blood cell production is maintained by hematopoietic stem cells (HSC) that self-renew as well as produce mature blood cells to meet normal physiologic requirements and respond to needs for increased production in response to stress or injury. HSC and lineage restricted progenitor cells (HPC) reside associated with bone marrow microenvirons or niches where non-hematopoietic stromal cells play key roles in determining their retention, fate, trafficking, and function. While hematopoietic microenvironments supporting stem cell function have been known for 40 years, specific populations of cells that contribute to HSC/HPC function have only recently begun to be identified, and controversy exists regarding niche composition and cell contribution. Despite advances, little is still known about cellular or molecular pathways contributing to niche regulation, about regulatory functions within or between niches, and how niche cells differentially regulate HSC functions under homeostasis or in response to stress or disease. We have had a long standing interest in the roles of prostaglandin E2 (PGE2) and the cyclooxygenase (COX) pathway on hematopoiesis and have made numerous conceptual advancements to the understanding of how PGE2 regulates normal and abnormal hematopoiesis. Our recent demonstration that PGE2 affects HSC function through regulation of homing and self-renewal and identification of molecular pathways and agents to fine tuning these effects, has facilitated current clinical testing of PGE2 to enhance HSC transplant. We recently found that blocking PGE2 signaling through the EP4 receptor using non-steroidal anti-inflammatory drugs (NSAIDs) results in differential HPC and HSC trafficking, particularly of HSC with enhanced engraftment potential. This finding is significant in that it highlights the importance of the PGE2 /COX pathway in the niche on HSC/HPC function; identifies the EP4 signaling pathway as a probe to investigate differential trafficking and heterogeneity of niche function; and validates the niche as a pharmacologic target. We believe this regulatory signaling axis is worth further exploration, but more importantly we believe discovery efforts exploring PGE2-EP4 signaling within the niche offer the opportunity to help delineate specific niche components, shed light on a current controversial topic and offers potential new clinical translation opportunities. In this proposal we will utilize the EP4 receptor as a molecular probe t test the hypothesis that PGE2, signaling in one or more cellular niches/niche cells, critically and differentially regulates stromal microenvirons, influencing both the trafficking (retention, egress and function (homing, engraftment) of HSC and HPC at homeostasis and in response to acute and chronic stress. Understanding cellular and paracrine interactions within marrow niches involved in HSC and HPC trafficking and function has major implications for collection of cells for lifesaving hematopoietic transplantation and treatment of various cancers with myeloablative therapies. Since NSAIDs block signaling through all 4 EP receptors, understanding the precise role of EP4 signaling will very likely identify more selective and efficacious therapeutic strategies.